What Is Endcapping in HPLC Columns

Endcapping the bonded phase of an HPLC column reduces silanol interactions and safeguards the silica support from dissolution. Various endcapping techniques are employed in modern HPLC column phases. Endcapping involves replacing the free silanol groups on a bonded stationary phase with trimethylsilyl (TMS) groups. The figure below illustrates the traditional stationary phase bonding and the TMS endcapping reaction:

What Is Double Endcapping?

Double endcapping means it is endcapped twice. Double (or triple) endcapping further minimizes silanol interactions and improves stability at mid pH.

When developing methods in the mid pH range, it is recommended to use an endcapped column. As pH increases, silica dissolution is decreased by endcapping.

When Would You Use a Non-Endcapped HPLC Phase?
Match your method’s pH range to the bonded phase pH Range.

Agilent Zorbax StableBond bonding is superior at low pH and stable down to pH 1. This is a non-endcapped bonded phase and is designed for selectivity and lifetime at a low pH. Shown below is an image representing the patented sterically protecting bonding, which is offered in 5 different selectivities (R= C18, C8, CN, Phenyl, or C3).

Agilent StableBond Reaction To Make a Sterically Protected Surface:

HPLC Columns for LC/MS

When choosing HPLC columns for LC/MS, chromatographers need to consider several aspects of their method and separation, typically including resolution, flow rate, and stationary phase choice. Often, for relatively simple analytes, shorter high-resolution columns are the best choice. These columns allow for high throughput while maintaining high separation efficiency.

Since many LC/MS analyses are run at lower flow rates (typically from µL/min flow rates up to 1 mL/min), moving to smaller internal diameter columns is the best choice for the user. Narrow bore (2.1 mm id) will often result in lower solvent usage for the method and provides better options for high resolution and higher sensitivity than the larger id columns. Most often, the best bonded phase choice is an endcapped C18 phase. For fast, high-throughput separations with LC/MS, the Agilent Poroshell 120 EC-C18 is an excellent choice. Poroshell has a larger frit, so it’s well suited for dirtier LC/MS samples, such as blood plasma, which may often clog columns with smaller porosity frits.

Conclusion

In order to extend column lifetime, ensure the method pH is in the bonded phase recommended pH range. In general, endcapped HPLC columns are a good choice for mid-range pH methods. Non-endcapped HPLC columns are a good choice for low pH methods.

The use of HPLC Syringe Filters, inline filters, and HPLC Guard columns are other ways to protect your analytical HPLC column. Please contact Chrom Tech for an HPLC Column recommendation or other HPLC consumables, including Idex Fittings and HPLC In-line filters.