Find out some of the most exciting ion chromatography applications, as reported by users, and more
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Ion chromatography (IC), a form of high-performance liquid chromatography (HPLC), is a process by which the anions and cation in an ionic solution can be separated and analyzed. Almost any charged molecules, including proteins and nucleotides, can be analyzed using IC. It is a broadly applied technique that is especially useful for clinical, industrial, food, and environmental samples.
Water Testing
Environmental analysis is one of the most important applications for IC. The technique can be used in the qualitative and quantitative analysis of anions and cations in drinking, surface, and saline waters, domestic and industrial waste, and leachates. Cyanide, hexavalent chromium, and bromate are examples of chemicals of interest in water testing with IC.
Carbohydrate Determination
Ion chromatography is increasingly being applied to carbohydrate determination in food science. Though they are not usually considered ionic, carbohydrates are acidic enough to be ionized in strongly basic solution, and are thus amenable to analysis by IC.
Pharmaceutical Analysis
Ion chromatography is a versatile tool for pharmaceutical analysis. The technique can be used to determine active ingredients, excipients, and traces of impurities, as well as metabolites in the form of organic and inorganic ions or polar substances, in certain drugs, drug solutions, and body fluids.
Ion exchange chromatography (IEX) complements the more common size exclusion chromatography (SEC) by offering a different approach to sample separation. While SEC focuses on separating molecules based on their hydrodynamic volume, IEX uses ionic charge as its separation principle. This combination allows for a more comprehensive understanding of samples by providing insights into size, molecular weight, and charge variation.
With IEX, a change in salt concentration and/or pH of the mobile phase during the experiment is used to elute samples off a column with a charged stationary phase. For example, with a stationary phase of negatively charged beads and a mobile phase of increasing salt composition, negatively charged species will elute earlier. Once the salt composition has increased sufficiently, positively charged species will elute. Coupled with MALS, information such as molecular weight and radius can be measured while simultaneously gleaning additional charge information from IEX. This strategy can be particularly beneficial in cases where different sample species have the same or similar hydrodynamic volume but different net charges, enabling separation with IEX.
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If you haven’t already set up HPLC CONNECT 4 and an ASTRA method for your system, doing so is easy! These topics are covered in this article: How do I use the HPLC CONNECT 4 software to control my Waters LC system? Open your ASTRA method within ASTRA via File | New | Experiment from Method.
Open an ASTRA Method file, select the Experiments icon, expand the Configuration option, and double-click the HPLC Instrument. This selection will display all of the information communicated to your Waters LC system to control its various modules, including your pump. In this window, click Edit Device Parameters to change parameters specific to your Waters LC system. In the pump method section, the pump time table can be changed to pull solutions of different salt compositions at different times during the method. In the example shown to the right, the salt content of the mobile phase has increased from the initial start to the end of the run.
When you have set up the composition profile for your IEX method, click OK to save these settings. Then, right-click the experiment name and select Save as Method to save this as a new method. Make sure you have set up your solvent reservoirs with the appropriate solutions! You are now ready for your IEX-MALS experiments.
Analysis within ASTRA for IEX-MALS experiments is similar to that for typical SEC-MALS experiments. Depending on your salt gradient, you may see a changing RI baseline through the run from your Optilab™ instrument. This is normal and expected, as the Optilab instrument will see changes in the concentration of your sample and the composition of the mobile phase. Run a blank injection, collect the data, and perform a Blank Baseline Subtraction and accounting for this shifting baseline. For more information, see our technical note TN on this topic.
If you regularly work with large solvent gradients, the Optilab High Concentration instrument may be better suited for your IEX-MALS needs. For example, the standard Optilab instrument can measure gradients between 0 – 450 mM NaCl without saturating, whereas the high concentration model can measure gradients up to 3.4 M NaCl.
Support for Waters Arc™ Premier HPLC control, including setting up methods for ion-exchange chromatography, is only available in HPLC CONNECT 4 with ASTRA 8.2.1 and higher. Future releases will add support for more Waters LC systems. For more details see the ASTRA 8 User’s Guide or the HPLC CONNECT 4 User’s Guide.
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