Filtering Samples for HPLC: More Than Just Column Protection

High-Performance Liquid Chromatography (HPLC) is a complex process requiring multiple steps that all need to work seamlessly together to get accurate and repeatable results from your samples. The pressure on analytical labs to process more samples in less time on the same equipment can mean that, not only are many analytical labs processing hundreds of samples in a week, but they are also often working at close to their maximum capacity. Any unexpected downtime is hard to recover. Even for analytical operations within smaller labs, which may not be running all the time, there are still concerns about getting the best data possible from the samples, and not wasting time on repeats, or on resetting an instrument because the column is no longer viable, or worse still, there are problems with the pump, the injector or other hardware requiring more lengthy and costly maintenance.

Why pre-filter samples for HPLC?

This concern over the performance of the instrument and the quality of the data has led to caution on the part of operators. One of the precautionary steps in widespread use is sample pre-filtration, where prior to loading the sample onto the instrument it is filtered, typically using a syringe filter, to ensure it is free of any particulate matter that might damage the injector or foul the column.

Particulate contamination in HPLC

Particulate damage can affect the HPLC instrument in several areas.  Two parts of the system that are most sensitive to particulate contamination are the injector where the sample must pass under pressure through a very narrow capillary, and the column.

Column occlusion

The problem most people are familiar with as a result of particulate contamination in HPLC, is blockage of the column frit by suspended particles resulting in increased pressure and a degradation in data quality. This problem is at least relatively easy to diagnose and can be fixed by changing the column. The downside, of course, is the cost of replacing the column, and the downtime on the instrument while this maintenance is performed.

Injector damage

Mechanical problems with the injector can arise as a result of particulate buildup.  Damaged injectors with leaks, worn seals, or capillary blockages can be relatively simple to diagnose, if still a lengthier task to fix than a damaged column. More troublesome can be intermittent faults resulting in unreliable sample injection that can remain below the threshold of detection with some samples failing and others not, causing costly and time-consuming repeat sample runs.

Guard columns

Guard columns and similar measures are employed on some systems to protect the analytical column. While these are a good idea, they are both expensive and hard-wired into the system which means they cannot easily be changed if they become blocked. Even samples with a relatively low level of particulate contamination can quickly occlude and overwhelm a guard column if large numbers of samples are being processed at a time. As guard columns become blocked by an accumulation of particles the pressure in the column will increase and peaks will become broader and possibly split, reducing the quality of data. Extending the life of these disposable, protective parts of the HPLC set up extends the service interval, reduces instrument downtime, and saves money.

Sample filtration extends the life of critical HPLC instrument components

Given all the problems that can creep in with even very low levels of sample contamination, pre-filtering samples prior to loading is a simple, rapid and inexpensive step that can greatly improve HPLC results. Sample pre-filtration represents a low-cost insurance policy for HPLC, extending the operational life of the expensive and critical to quality instrument components including pump, injector, and column.

Filter selection is critical to success. Using the wrong filter membrane can result in contamination or other damage to the sample with a negative effect on the results. Different mobile phases require different filter membranes to ensure that undesirable materials are not being leached out of the membrane and into the sample. Different samples also require different membranes to ensure that analytes are not bound by the membrane and removed or reduced from the sample prior to injection.

In our next blog, we review the filter selection process, and how to choose the best filter for your setup, optimizing your analytical protocol to improve sample data and extend the life of the components in your HPLC system.

As well as producing a wide range of filters suitable for sample and mobile phase filtration in any HPLC application, our website also contains some excellent resources to help you better understand pre-filtration in analytical protocols and to select the best filters for your needs.