Ion Chromatography - Chemistry LibreTexts

This instructor’s guide includes sample answers to the questions in the IC module, and additional information at times.

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I have given this module as a take home assignment for the students to work on and read through, then had them present their answers to each other in small groups. This resulted in the students having discussion on where their answers were different and did well in getting to the underlying principles.

Q1. Write the chemical reaction for the association of A- and B- with the ion exchange resin.

\[\ce{Resin^+-E- + A- <=> Resin^+-A- + E- }\nonumber\]

\[\ce{Resin^+-E- + B- <=> Resin^+-B- + E- }\nonumber\]

Q2. Write the chemical reaction for the elution of A- and B- from the ion exchange resin.

\[\ce{Resin^+-A- + E- <=> Resin^+-E- + A- }\nonumber\]

\[\ce{Resin^+-B- + E- <=> Resin^+-E- + B- }\nonumber\]

Q3. Write the equilibrium expression constant for A- and B-. In chromatographic separations, this term (Kc) is referred to as the distribution coefficient.

Q4. Do you think the magnitude of the distribution coefficients are the same for A- and B-? Why or why not?

If they are different ions, then they would have different distribution constants, because they would have different affinities for the resins. These differences will arise due to differences in size, charge, and other characteristics.

Q5. If the distribution coefficient of A-() is smaller than the distribution coefficient of B-(), draw a sketch of the elution process that is similar to the figure in Step 1 but at a point where A- and B- are partway through the column.

Q6. Suppose B- had a very strong affinity for the resin, what would happen to its elution time?

As the affinity for the resin increases the elution time increases. Thus you could get to a point where the elution time is so long it does not appear to come off the column.

As the sample is injected onto the column, the two different analytes briefly displace the eluent as the counter-ion to the charged resin. The analyte is briefly retained at the fixed charge on the resin surface. The analytes are subsequently displaced by the eluent ions as the eluent is added to the column. The different affinities (see the chemical reactions in the basic process section) are the basis for the separation. The Kf value of each reaction is also known as the selectivity coefficient. The greater the difference between the Kf values for the two analytes, the more the two analytes will be separated during the ion chromatography process. In reality, the interaction between the solvent and the analyte can also have an impact on the order each analyte is eluted. For a more in-depth analysis of predicting the retention order see the material by Dr. Thomas Wenzel. (http://www.bates.edu/x.xml)

Q7. Is this a desirable or undesirable situation if you were trying to analyze A- and B- in a mixture?

You want have a difference in elution times, but you do not want the affinity to be so strong that the ion does not readily come off the column. The longer the elution time the more peak broadening will happen (this links to later brief information on peak broadening.)

Q8. Is there a situation you can think of when it might be desirable for B- to have a very strong affinity for the resin?

Water filtration systems, such as water softeners, usually have an ion exchange resin where the affinities are strong enough to effectively remove ions from the water. The Mg2+ or Ca2+ displaces the Na+ on the resin. The affinity for the Mg2+ or Ca2+ ions needs to be much stronger than the affinity for Na+ in order to effectively soften the water. In sample pretreatment you may want very strong affinities to bind all of the ions and then extract with a different solvent.

Q9. If you want to separate cations, what would be different about the stationary and mobile phases?

The resin would need to be negatively charged, and the mobile phase would be positively charged. The common cation exchange resins are based on either polystyrene-divinylbenzene (PS-DVB) or methacrylate polymers. The surface of these polymers (Figure 1) is functionalized with a negatively charged sulfonated group (-SO3- ). The cation in the eluent or the analyte of interest is the counter-ion in the vicinity of the charged functional group.

The surface of the polymer is functionalized with a quaternary amine (-N+R3) for anion exchange (see Figure 2). The quaternary amine provides a positive charge to the surface, attracting negatively charged anions in the liquid phase. Just like the cation exchange resin, the anion of the eluent or the analyte of interest exists as the counter-ion in the vicinity of the positive charge residing on the amine.

Q10. Would water flow easily through a column containing very fine particles?

As the resin particle become finer, they pack together closer, leaving less room for the solution to flow through. This, in conjunction with the more resin-solvent interactions, will increase the resistance to flow. You then have to apply pressure to force the solutions through the column.

Q11. If not, how could you get the water through the column?

As the water becomes less likely to flow, you need to apply a force to push the water through the column. This is done with a pump that can handle higher pressures (often 200- psi) such as a double piston high pressure pump to force the mobile phase through the column.

Q12. Considering that the column is packed with very fine particles, what must be done to surface water samples before injecting them onto the column?

If the column particles are very fine, the surface water sample will likely need filtered to remove any detritus that would foul up the column. Surface water samples are filtered through at least 0.45 µm filters but may be filtered through as small as a 0.20 µm, similar to how other solutions are filtered for use in IC. The filter type is one that must not introduce an error.

Q13. Can you think of a way to detect the presence of ionic substances in water?

The most common factor to all ionic substances is that they have a charge, thus a detection method that is based on measuring the charge is frequently used. Ions in solution conduct electricity, so the conductivity of a solution will change as the concentration of ions change. This is the common basic detection. There are other possibilities that are more selective.

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Conductivity would not distinguish between the two ions. Therefore you need the two ions to be separated on the column.

The selectivity in the overall method comes from a good separation of the ions, so the detection method does not need to be very selective. The detection does need to be quantifiable and applicable to a broad range of ions.

Q16. Will the eluent ion respond to the detection method you thought of above to measure the presence of Na+ and Ca2+.

Yes it will. Thus eluent suppression will usually be used.

Q17. How will the chromatogram change as you increase the concentration of A- and B- injected into the column? Make sure to label the axes of your chromatogram.

x-axis is elution time, y-axis is conductivity. As you increase the concentration of an ion, the height and area of the peak corresponding to that ion will increase.

Q18. Since neither axis in the chromatogram you drew above is concentration, how can we calibrate the detector response to determine the concentration of A- and B-?

The analysis of beverages is extremely important for the general health of the population. Why is this so? Our bodies are composed of about 60% water, depending on several factors like weight and sex. Hydration is one of our basic physiological needs, as noted in Maslow’s hierarchy of needs. When we are dehydrated, a number of problems occur, from irritability to confusion leading to severe kidney problems and even low blood volume shock in extreme cases. Therefore it is incredibly important for standards to be set by regulatory agencies regarding the contents of the beverages we choose to drink, whether this is water, milk, coffee, juice, soft drinks, beer, wine, or any other number of items. Reliable beverage analysis is critical for many reasons: product monitoring and quality control, general content determination, and to avoid health issues.

Ion chromatography is a simple and robust analysis technique that is able to measure several components in beverages with relative ease compared to these other technologies.

Typically, conductivity detection is used for IC analysis. Other options are available including UV/VIS and amperometric detectors for more specialized analyses (e.g., carbohydrate analysis).

Find out more about our comprehensive options for IC detection

When analyzing complex beverage matrices like milk, coffee, or wine, sample preparation steps are normally required to protect the instrument (e.g., from contamination or blockages due to particles). Performing these steps manually is a very time-consuming and costly process that is also prone to human errors. Metrohm offers a time-saving solution for this with «MISP»: Metrohm Inline Sample Preparation specifically developed for difficult sample matrices. Several options are available including Inline Ultrafiltration, Inline Dialysis, Inline Dilution, and much more.

Watch our LabCast video below about to learn more about the benefits of using Inline Ultrafiltration in IC.

FAQ about beverage analysis with IC

Now that you know a bit more about the capabilities of ion chromatography for quality control in beverage analysis, it’s time to answer some frequently asked questions in this field. Dr. Gabriele Zierfels, Senior Product Specialist Ion Chromatography at Metrohm, has given a webinar hosted by New Food Magazine discussing how IC can help modern quality control labs from the beverage industry comply with official quality and labelling standards and make their daily routine analytics more efficient, which you can watch on-demand for free.

The webinar begins with an overview of the latest analytical techniques used by the beverage industry to comply with quality standards and labelling requirements such as EU regulation / and US regulation 21CFR101. Then the focus shifts to the versatility of ion chromatography for beverage testing and how it can help modern QC labs increase the efficiency of their daily routine analytics, which is exemplified in the main part of the webinar by numerous application examples.

Here we answer the top five questions asked by participants regarding beverage analysis with ion chromatography after the webinar.

Webinar: Improving your analytics for beverage analysis: How ion chromatography can help the modern QC lab become more efficient

1. What are the differences between HPLC and ion chromatography (IC) when it comes to beverage analysis? What are the main benefits of using IC?

High-performance liquid chromatography (HPLC) is typically used to separate complex mixtures with large organic (nonpolar) molecules by utilizing their affinities for different solvents and interactions with modified stationary phases. Many analytes required for food and beverage testing are either ions or polar molecules, some of which cannot be measured using reversed-phase HPLC.

IC on the other hand is a simple and robust analysis technique which allows determination of similar chemical substances in a single chromatographic run. With IC, ionic or polar analytes can be determined in very complex matrices with superior sensitivity and reproducibility using analytical separation columns made of ion exchange resins. The analytes undergo chemical/electrostatic interactions with the column resin. Due to such interactions these analytes are retained stronger than on reversed-phase columns. This allows excellent separation from the matrix components.

Check out the benefits of using IC over HPLC in our video.

2. How easy or difficult is it to switch between applications with a single instrument setup (e.g., analyzing different beverages like coffee and juices)?

Switching between different sample types can be simple, but every sample requires preparation before injection into the chromatographic system. In most cases this means sample dilution or filtration. This procedure can be done manually (which is time consuming) or completely unattended utilizing automated Metrohm Inline Sample Preparation (MISP) techniques. Therefore, several different sample types (e.g., tea, coffee, or juices) can be analyzed one after another for the same analyte profile, such as sugar content. The sample matrices can vary widely, as Metrohm offers various MISP techniques to get the cleanest possible extract for injection and subsequent separation and quantification of the target analytes.

Download free Application Notes below to learn about the analysis of a variety of analytes in multiple beverage types with IC.

IC applications for drinking water

IC applications for nonalcoholic beverages

IC applications for alcoholic beverages

3. How robust is IC when it comes to analytes that require stabilization, for example, sulfite?

Determination of samples containing analytes that must be stabilized prior to analysis (e.g., sulfite) is even more robust when using IC for the task. Even if samples have been stabilized, the detection can be disturbed by electrode fouling in the amperometric detector. To avoid this process (which is common in Direct Current mode), a short automatic cleaning method was applied between the sample analyses for stabilization of the signal and results that lasts for up to three weeks. This means no manual polishing steps and no disposable accessories are required.

To learn more about simplified sulfite analysis with Metrohm ion chromatography, download our free White Paper, check out our previous blog post, and download our free article featured in LC/GC’s The Column.

White Paper: Simplified sulfite determination in foods and beverages with ion chromatography

«Analyze This»: Determining the total sulfite in food and beverages: faster and easier than ever

LC/GC article: A Simplified Method to Determine Total Sulphite Content in Food and Beverages via Ion Chromatography

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