HPLC column protection
Use of high-performance liquid chromatography (HPLC) to characterize samples is an invaluable technique across numerous industries including the testing of pharmaceuticals, foods, and environmental samples among others.
Because HPLC columns are central equipment in many laboratories, protecting them from damage is a prime consideration to increase accuracy of test results. A well-maintained column also produces consistent and accurate data, supporting confident analyses.
Increase Column Performance
Column performance depends on several factors, including sample properties, presence of insoluble particles, and cleaning and maintenance procedures.
It is not always feasible to work with the most favorable conditions for column lifetime. For example, some samples have harsh but necessary pH conditions. However, appropriate column protection techniques and preventative maintenance procedures can help to maximize the life of a column and maintain HPLC efficiency.
What factors affect column lifetime?
Commonly reported HPLC problems include slow flow rate and increased column back pressure. Either of these observations could indicate that a column’s performance is being compromised.
Particulates from unfiltered samples or mobile phases can contribute to these issues. Essentially, anything that enters the HPLC system runs the risk of introducing particulate matter onto the column matrix, potentially shortening the useful column lifetime.
Several other factors can cause damage to the bonded phase in a column, reducing its efficacy. These factors include reactive material in samples, harsh pH conditions, and excessive pressure. Chemical incompatibility between sample solvent and sample preparation consumables can also be damaging.
How can I protect the integrity of my results?
1. Sample preparation
Prefiltering or centrifuging samples prior to injection onto the column can help to limit the addition of particulates to the system. A syringe filter or syringeless filter vial is suitable for this step, depending on the sample type, number of samples, and sample injection method.
Chemical based sample pretreatment is another way to protect column performance. Many different techniques are possible. The potential issue here is expense and time, calling for an appropriate balance between additional pretreatment costs and their possible cost-saving benefits.
2. Mobile phase
The mobile phase is another factor that influences column performance.
Filtering and degassing the mobile phase will remove particulates and air, ensuring they do not reach and damage the column. Filtration is especially important if the mobile phase contains buffer salts or modifiers from dissolved solid material.
Also, it can be useful to consider the compatibility of a sample with the mobile phase, because some sample components are reactive with particular HPLC solvents. For example, aldehydes and ketones react with an amino mobile phase. Avoiding reactive combinations helps to protect columns against damage.
3. In-line filters and guard columns
In-line filters and guard columns help prevent particulates entering the system and the column. The use of these column protectors depends on the application, with either one, or both, required.
A guard column is a short column that the sample and mobile phase pass through before entering the main column, helping to protect against damage by adsorbing particulates. The use of a guard column depends on the specific application, because it can be an expensive protection option requiring consideration of the costs against benefits.
Maintaining a clean column is another way to extend a column’s lifetime. Occasionally flushing the column with mobile phases that differ from those used on the system (e.g., more water or organics) can help to clean the column. Extensive flushing in a backwards direction can also help with cleaning, especially if you notice increasing column back pressure.
Storing columns in an appropriate solvent can help prolong their life Always consult the column’s handling instructions to identify the best storage solvent for that particular column.
Try our Whatman Filter Selector App to find out if you are using the most appropriate filtration solution for your samples. To discuss any challenges you are facing, please contact GE's Life Sciences Scientific Support.