Tips & Tricks GPC/SEC: How to Care for your GPC/SEC Instrument

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The Column

Column, The Column-10-05-2015, Volume 11, Issue 18

Tips & Tricks GPC/SEC: How to Care for your GPC/SEC Instrument

Columns are at the heart of a gel permeation/size-exclusion chromatography (GPC/SEC) system. Software is used to evaluate the data, to understand and present the results, and to act as an interface between the analyst and the system. But what about GPC/SEC pumps, injection systems, and detectors? They should run uninterrupted 24/7, without the analyst having to think about them or to deal with them, but these components also need care. Following some simple rules can give better results in less time. 

  Modern gel permeation/size-exclusion chromatography GPC/SEC pumps, injection systems, and detectors are (in general) very robust and stable against lots of different solvents/mobile phases. Some precautions are required, however, in daily operation, even when using the best components. Nearly all of these precautions are associated with the mobile phase that is applied.   

Mobile Phase Selection

All solvents should be of the highest quality (HPLC-grade). Even if these reagents are more costly, the difference in purity is marked. It is very important that the mobile phase is free of particles and dust because this might otherwise cause blockages in the system or the columns.   It is good practice before using a new solvent to verify that all parts of the GPC/SEC system are compatible with the solvent and, when using aqueous systems, can be used at the pH value applied. This is especially important if components are used that have originally been designed for high performance liquid chromatography (HPLC). Unfortunately, GPC/SEC analysis often requires “exotic” organic GPC/SEC mobile phases that might cause problems with seals or other parts of the components.    If the application allows, the preferred choice of solvent should be a non-corrosive solvent. For example, if an application can be run in tetrahydrofuran (THF) or in chloroform, from an instrument point of view the choice should be THF.    Many GPC/SEC applications require the addition of salt, acidic, or basic additives. The concentration of these additives should always be kept as low as possible. In the case of dimethylacetamide (DMAc) or dimethylformamide (DMF), lithium (Li) salts are often used as additives. Because of the increased solubility of LiBr compared to LiCl, bromide is recommended in many application notes. However, it is more corrosive than LiCl and therefore potentially more dangerous for stainless steel instrumentation.    In aqueous systems, the addition of an additive preventing algae growth is recommended. Adding 0.05% NaN


or acetonitrile will suppress the growth of microorganisms that could otherwise block tubing or columns.    Seal wash options should be installed if mobile phases with salts are used. The best seal wash liquid is in most cases the pure solvent. Seal wash liquids should also be exchanged regularly and if the seal wash seals and holders are installed it is important that they are not running dry.    If very high salt concentrations or extreme pH values are required it might be a good investment to use stainless-steel free instrumentation, which is now commercially available in many flavours and for nearly all detection options.   

Preventive Maintenance 

Preventive maintenance should be performed regularly to avoid unexpected instrument downtime, to ensure highest data quality, and to avoid costly repairs as a result of secondary damages as a consequence of not replacing worn parts.    In every GPC/SEC system there are at least some in-line filters, pump seals, and injection rotor seals that require frequent exchange. In addition, check valves, pistons, tubing, and lamps should be checked regularly and replaced if required.    The cycle for replacing of worn parts depends again on the application. As a rule of thumb we can say that one preventive routine maintenance a year is sufficient for a system with salt-free applications that is running regularly. If the application is more demanding and corrosive solutions are used, a reduced cycle of 6 months (or even less) is recommended. 


Idle Mode, Weekend, and Vacation 

Users sometimes have to decide if it is worth powering down the GPC/SEC system or not.    Reducing the flow-rate is a good option that helps to save mobile phase, but still allows the instrument to be started up again very quickly.    For a short shutdown period of a few days or less, it is also possible to run the system in recycle mode. In this case, the effluent from the detector is redirected into the solvent reservoir. However, running a GPC/SEC system should only be done if no injections are performed and the mobile phase is free from salt and additives (with the exception of algae prevention additives). Even in the case of running the instrument in recycle mode, the mobile phase should be exchanged regularly.    If an instrument is not needed for several weeks or months it can be powered off completely.    Special care is required for instrumentation and columns when mobile-phases with salts or other additives are used. As long as there are salts or additives in the system, a low flow-rate should always be applied (at least 0.05–0.1 mL/min) to prevent corrosion of the instrument or the columns.    If the pump is going to be completely turned off, the salt solution should first be replaced by pure eluent. For this, at least 3–5 column volumes of pure mobile phase, if not more, should be used.    If the instrument is not used for a long time the separation columns can be detached and stored tightly plugged with their original plugs in the refrigerator (without freezing them). In the case of corrosive mobile phases (chloroform), the instrument can be switched to a different mobile phase (isopropanol) prior to shutdown. Restarting the GPC/SEC system then requires a switch back to the original mobile phase. For new sample runs the GPC/SEC mobile phases and buffers should be prepared freshly on the day required. Starting an analysis with solutions that are several days (or even weeks) old and have been run in recycle mode will most probably end with low quality data with drifting and wavy baselines.    If the columns were reinstalled it is good practice to first apply a low flow-rate (to ensure that there is no air from storage trapped) and to flush the columns with at least 3–5 column volumes before attaching them to the detectors. It is also worth checking the calibration with a checkout sample. If in doubt a new calibration should be performed.   


  • Preventive maintenance helps to increase instrument uptime.

  • GPC/SEC instruments always need a low flow-rate if mobile phases with additives or salt are used. They should only be turned off once removal of all salts and additives using the pure mobile phase has been performed.

  • Verification of solvent and pH compatibility is required particularly for “exotic” organic phases. Dedicated instrumentation for extreme pH values or applications with high salt content is available. 

Daniela Held

studied polymer chemistry in Mainz, Germany, and works in the PSS software and instrument department. She is also responsible for education and customer training.   E-mail: 



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