Most HPLC columns are made from stainless steel, the actual separation happens into the column usually columns are 30 to 250 mm in length, 01 to 05 mm in diameter, and 03, 05, and 10 microns in pore size. The column is packed with porous particles. The Porous particles are made from polymer and surrounded by a thin equal layer of silica and polystyrene.
In the early years, LC was carried out in a glass column with a diameter of 01 to 05 cm and a length of 50 to 500 cm. Commonly HPLC has a guard column in front of the analytical column to protect from contaminants and remove particulate material it helps to extend the life of the analytical column, the guard column and analytical column have the same stationary phase.
As far as I recognize, there may be no "best before" date. if you store it in the appropriate solvents and you preserve it in the right condition, I do think it may still work, however, the performance might not be exactly what it was. however, you'll have to test it out.
What I commonly endorse is to make a chromatogram of what you assume with the whole method and if you want to use it again in a year or 2 years, go back to those conditions and that actual method and try it out and take a look at the chromatogram now vs before. that will tell you how appropriate the column is still.
Obviously, 2 years is a very long time to have a column sitting on a shelf, so I would not be too surprised if the performance drops a little bit at least. If you want the least performance drop, I would suggest giving it a good cleaning. Then put it in appropriate solvents. Store it in a zone to avoid vibration, extreme temperature or moisture.
To ensure product quality, the columns should ideally be stored at ambient conditions (unless otherwise specified in care and use instructions) with sufficient controls to prevent adulteration and contamination. Under these conditions, columns can have an extended lifetime. However, because of the diversity of applications that can be associated with columns, it is the policy of column manufacturers to consign the task of expiration dating of the columns to the customer. Therefore, strongly recommends that the expiration of the column(s) be assigned by the customer based on his or her individual application.
Letting move of your favorite column may be difficult to do, so whilst is it without a doubt time to say goodbye? underneath are a number of the signs to look for that your courting and your HPLC column lifetime is coming to an end.
- High Backpressure not reduced by reverse flushing the column
- Split Peaks
- Loss of Resolution
- Broad Peak Shape
- Retention Time Shifts
High Backpressure
- Increases in backpressure are typically the result of a blockage at the inlet frit of the HPLC column. To remove the blockage in most situations you can reverse the flow of the solvent through the column at a reduced flow rate and flow 100% strong solvent through the column to remove this. If your pressure does not return to normal either the frit is blocked irreversibly or there is a void in your column. Both situations require the column to be replaced.
Many split peaks are also the result of a blocked frit at the inlet of the column which results in a channeled injection. To resolve this follow the steps outlined above first and foremost. If the split peak remains the likely cause is either incorrect running conditions of your method (working too close to the pKa of your compound or using a strong injection solvent) or your column may be fouled with strongly retained contaminants. If the inlet is irreversibly plugged or the stationary phase fouled with contaminants you will need to replace the column.
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Loss of Resolution
- HPLC columns are consumable items and as such over time you will begin to lose the resolution of your critical pairs. Some column regeneration may be possible and each column will have its own individual protocol for this, however, once a loss of resolution of a critical pair of compounds has been reached and cannot be regenerated, your column has reached the end of it’s natural life for the particular analysis you are doing.
Broad Peak Shape
- Some peak broadening can occur for reasons other than the natural end of the life of a column, these include
- Injection overload
- Inadequate buffering
- Inappropriate method conditions
- Strong injection solvents.
- However, if these elements have been eliminated, peak broadening is one sign your column is aging. Once your peak shape has deteriorated to the point you can no longer pass system suitability or resolve critical pairs it is time to part ways.
Retention Time Shifts
- More often than not retention time shifts are the result of poor method control, however, in the case of phase contamination, you will see retention time drifts of peaks if your phase is becoming fouled. Not all peaks will be affected equally and if you have eliminated all method concerns from your troubleshooting the likely cause is stationary phase contamination and your column will need to go.
To be certain the column is the problem there are a few things you can test
- Clean the column either by a reverse flush or other method outlined in the specific column care guide; each column has an individual guide but a general one for most columns can be found here.
What is the lifetime of an HPLC column if they are stored with endcaps?
- HPLC column can be fine for a long period of time and has no defined shelf life, but it is dependent on what storage solvent was used for column storage. If the column was properly flushed after use and stored in 100% organic solvent such as acetonitrile, the column will survive for many years.
- The best practice is to use the shipping solvent of the column as recommended by the column manufacturer. If the column was not flushed and stored under buffer conditions, for instance, the lifetime will be significantly reduced due to the precipitation of the buffer salts. The history of the column can have an influence on its lifetime of the column. It suggests running the QC test as described in the information supplied with the column.
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HPLC
