How Many Injections Can I Get on My HPLC Column?

It depends! Many factors affect how many injections you can get on your column, including:

• Injecting “dirty” samples
• Following HPLC Column manufacturers recommended protocol/operating conditions
• Using HPLC accessories, like guard columns or inline filters to protect your HPLC column

All of the above determines how many injections you can get on your HPLC Column.

What Are You Injecting on To Your HPLC Column?

When chromatographers have relatively clean samples vs. “dirty” samples, one should expect to get significantly more injections. Pharmaceutical companies analyzing product formulations or simple drug mixtures and standards have relatively clean samples and would expect to get thousands of injections on a single column. Clinical laboratories analyzing biological samples are typically analyzing “dirty” samples (even after sample preparation), and these labs would typically have fewer injections per column than those labs with clean samples.

Are You Following the HPLC Column Manufacturers Recommended Operating Conditions?

All HPLC columns have recommended operating conditions, including temperature limit, pH range, and an upper pressure limit. Operating outside of the recommended parameters will significantly reduce how long your column will last. For example, when using a column below the recommended pH range, it can cause hydrolysis and detachment of the bonded phase from the silica. Operating above the recommended pH range for your column could cause dissolution of the silica gel. Also, If the column is continuously operated at its upper pressure or temperature limit, it will negatively affect the lifetime and the number of injections on the column.

Which HPLC Column Phase Are You Using?

Some phases are inherently more rugged than other phases. Over time, the stationary phase backbone will start to breakdown, which forms voids in the column. Chromatographers will then notice peak splitting and peak broadening, which will result in a loss of resolution and sensitivity.

Are You Protecting Your HPLC Column?

Using HPLC accessories, like guard columns or inline filters, adds an additional level of protection to the HPLC column and is recommended to maximize the number of injections on the HPLC column. Inline filters are an inexpensive option but do not have any bonded phase, so the main purpose would be to trap particulates. Typically, guard columns are selected to match the analytical column’s phase. Therefore, if an analyte would irreversibly bind to the phase, it would be trapped on the guard (which then can be replaced as needed) instead of passing through and binding on the HPLC column.

Conclusion:

Since many factors affect how many injections one can get on an HPLC column, it is not possible to predict how many injections someone should get on an HPLC column for a new method. Modern HPLC systems have built-in column modules and software that monitor the injection number on the column. Using this counter can be an effective troubleshooting tool. If a specific application routinely gets 2,000 injections on an HPLC column and, all of a sudden, the same method/column only gets 500 injections, this will lead the chromatographer to troubleshoot the method to see if anything changed.

• Was a fresh mobile phase prepared (and made to the wrong pH)?
• Did a new batch of mobile phase solvent come in (and have contamination)?
• Did something change during the sample preparation procedure?
• Microbial Growth in Solvent Bottles (or other portions of the system)?

Please reach out to Chrom Tech to discuss which HPLC accessories are needed to maximize the number of injections on your HPLC Column.