Monday, 14 June 2010

Help on: Number of Injections for HPLC Columns


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"How many injections should I expect to get out of my reversed phase C18 HPLC columns?"
“Put simply, it depends. This is probably not the definitive answer you were hoping for but there are a number of different factors which influence the total number of injections that you can expect for a HPLC column. These relate to the actual column in question, the way you are using the column (the method operating conditions), and also the nature of the sample which is being injected.
Factors relating to the column include the type of packing, e.g. silica, zirconia, etc., and how the stationary phase, in this case C18, is bonded to the packing. Factors relating to the operating conditions include the pH, temperature, and mobile phase components such as solvents and buffers. Factors relating to the sample include the cleanliness of the sample, the volume, pH, impurities present and the nature of the actual sample molecules. ‘Clean’ samples are usually straightforward preparations such as standards, simple drug formulations and mixtures, whereas ‘dirty’ samples include biological fluids and environmental samples.
You can maximise the number of injections for a column by using it within its recommended operating conditions, e.g. pH and flow rate, and by flushing the column routinely to remove strongly retained impurities. Use of a guard column should also help to prolong column life.
As a rough guide you can expect to achieve at least 1000 injections if the column is used within its recommended range of operating conditions and the sample being injected is ‘clean’. You may even be able to perform 5000 injections for some methods. If using ‘dirty’ samples then you should expect to achieve less than 1000 injections but the actual number can vary considerably and may even be as low as 50 injections.
Of course if you do not count the number of injections performed on your columns then you will not know how many injections are possible. It is helpful to keep a record, if only for budgeting purposes in these lean times. Modern chromatography data systems and autosamplers make it simple to count injections for a particular column as long as you keep a note of a column identifier such as serial number.”

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