15th May 2024
Optimizing HPLC Methods with Agilent’s InfinityLab Poroshell 120 Columns
More Chemistries, More Choices for Separation
The InfinityLab Poroshell 120 family has grown to include three particle sizes and 20 chemistries, so you can efficiently separate the widest variety of compounds. But, with all these options, how do you know what column to use for your method? Whether you’re in HPLC method development or looking to optimize your existing method, this guide will help you determine what column is best for your application.
Understanding the Importance of Column Particle Size
Selecting an appropriate particle size for an HPLC column is crucial for optimizing your method. The particle size affects the efficiency and resolution of the separation process. Agilent’s InfinityLab Poroshell columns are available in 1.9 μm, 2.7 μm, and 4 μm. Each size has its own advantages and considerations.
The smaller the particle size, the higher efficiency and better resolution without lengthening the column or increasing run time. However, it will increase the backpressure, which may require the use of a UHPLC system (as opposed to a standard HPLC system) to accommodate. Larger particle sizes have lower backpressure and may provide sufficient resolution for various applications.
It is important to consider the specific method and HPLC instrument requirements while choosing a particle size. See below for a general guideline:
Figure 1: Particle Size Guideline
Choosing the Right Column ID and Length for your HPLC Method
In addition to particle size, selecting the right column ID (inner diameter) and length is also essential for optimizing your HPLC method.
The column ID affects sensitivity and the amount of solvent required for the separation. Legacy methods typically use a larger column ID (e.g. 4.6 mm), but often a smaller ID is preferred for higher resolution and better peak shape.
The column length affects the separation efficiency and the resolution of the method. Shorter columns, between 30 to 100 mm are used for fast separations, whereas longer columns greater than 150 mm are used for increased resolution.
Comparing Poroshell Chemistries to Zorbax Chemistry-Based Methods
There are many benefits to switching from fully porous columns to superficially porous columns. We dive into this in greater detail in another blog post, but for this guide, we’ll simply jump into method transfer from a fully porous to superficially porous particle column. If you’re already using an Agilent ZORBAX column, making the switch to an InfinityLab Poroshell column will be easy. The InfinityLab Poroshell chemistries are aligned with the traditional ZORBAX chemistries. See below for more info:
Figure 2: Switching from ZORBAX to InfinityLab Poroshell columns
When transferring from Zorbax to Poroshell columns, consider the particle size, column ID, and length that were previously used. It may be necessary to optimize the method parameters slightly to achieve the best results.
Exploring the Variety of Poroshell Chemistries Available
Agilent offers a variety of Poroshell chemistries to suit different separation needs. Some of the most commonly used InfinityLab Poroshell chemistries include:
Purpose
Stablebond C8 & C18
Endcapped C8 & C18
& Low pH
Selectivity
for Unique Selectivities
These are just a few examples of the chemistries available. For more specific and detailed information, I recommend checking out Agilent’s InfinityLab Poroshell column guide. The comprehensive range of columns ensures you can find the right chemistry for your application.
- Determine the optimal particle size, column ID, and column length for your HPLC and application
- Optimize the mobile phase composition, flow rate, and temperature to achieve the best separation and resolution
- Take advantage of Chrom Tech’s technical support and resources for any troubleshooting or method development assistance.
By following these tips, you can maximize the performance of your HPLC method and achieve reliable and reproducible results.
