![]() Preferably it should be replaced as a regular preventative maintenance procedure before adverse effects are noticed. Guard cartridge is saturated with matrix- This should be replaced any time a change in peak shape or retention is noticed.Changes in instrument settings- see discussion above for this.Error in preparation of mobile phases- It is always helpful to carefully remake the mobile phases to rule this out.Sample volume too high, overloaded- Peaks usually are fronting under these conditions, but sometimes can tail.Try making your sample in your mobile phase solvents at the same ratio as the starting conditions of your gradient (or same ratio as your mobile phases if using isocratic conditions). Solvent composition of sample is not compatible with the mobile phase or gradient- This is usually accompanied by poor peak shape.This could be caused by any of the following: If early eluting peaks are shifted, but late eluting peaks are relatively unaffected, the issue is usually related to differences in the solvent ratio with sample and/or mobile phase. Please see the following post to find out which columns are compatible and which are not.Ĭan I use a 100% aqueous mobile phase with my LC column? This is caused by an attempt to use a mobile phase that is more than 95% water/aqueous with a column that is not aqueous compatible. If all peaks are dramatically shifted to an early retention time, coming out together at or near the void volume, the issue is likely phase dewetting, or what we used to call phase collapse. Make sure you have the column in a thermostatic column compartment to avoid this. Changes in column temperature- this can affect the column in a way similar to flow rate and can move the retention in either direction.When multiple users are performing the same analysis on shared equipment, this can often be an issue. Changes in instrument settings- This may seem obvious, but one should always confirm that program parameters have not changed from time to time.Filtration systems such as this also serve to degas mobile phases because the solvent is pulled through by vacuum. We have a membrane microfiltration system available for this if needed. Please note that any mobile phases prepared to contain buffer salts or any modifiers in solid form should be filtered prior to use for LC. If a degasser is not available, mobile phase can be degassed manually before using for LC. Bubbles in the pump or tubing- Make sure your system has a degasser built into the system that degasses solvent prior to pulling it into the solvent pump.Faulty check valves- Get help from an instrument service person if needed to verify performance of check valves and other parts in the flow path.Worn or faulty pump seals- Replacing the seals is often a good precaution to prevent this.Leaks in the flow path- Check to make sure all fittings are tight and in good condition.Flow rate variations can be caused by any of the following: This can be verified by collecting eluent from the column into a graduated cylinder and using a stopwatch to measure the time taken to fill the container to the mark. This can be a shift in either direction, but is often toward longer retention times/slower flow rate. If all of the peaks are shifted by about the same time interval, the variations are likely caused by variation in flow rate in the instrument. An important factor is whether all of your peaks have shifted or whether it is only certain ones. It is always useful to define the specific problem to help narrow down the possible causes. Also we are assuming that the analyst is following an established method that worked for a significant period of time prior to seeing changes in retention. For the purposes of this discussion, we will be discussing changes in retention on one individual column, not differences from one column to another. Hopefully this post will help you navigate through troubleshooting this situation. If you have experienced retention time shifts with LC, you know it can be complicated and can cause problems with quantitation if not resolved. ASE Extraction System Parts Cross-Reference.Documentation Search (SDS, Certs, Data Packs).EZGC Method Translator & Flow Calculator. ![]()
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