Aeris Core-Shell LC Columns for Proteins and Peptides

The world is changing and we need to widen our Environmental Scope. Although analytical techniques and established regulated methods are not easily changed; more complex targets, larger lists, and a variety of sample matrices create a daily challenge for environmental chemists. By using the latest technologies and techniques in HPLC/ UHPLC and Sample Preparation, laboratories and chemists are able to develop better approaches to make their work Faster, Cheaper and Easier. Phenomenex is committed to supporting the industry through a comprehensive product portfolio of Sample Preparation, HPLC/UHPLC, and LC-MS, columns and accessories, along with application and method development services to meet industry needs.

Cyclosporine A, tacrolimus, sirolimus, and everolimus are four of the most commonly administered immunosuppressant drugs and play a central role in the success of tissue and organ transplants. These drugs are most typically analyzed from whole blood using LC/MS/MS but this presents analytical challenges when working with whole blood. This application note presents a rapid and effective method for the analysis of these four immunosuppressants from whole blood that use a simple protein precipitation step followed by direct injection onto a wide-pore core-shell HPLC column (Aeris WIDEPORE 3.6 μm XB-C18).

This application note demonstrates the use of Aeris™ WIDEPORE core-shell columns in the improved resolution in large proteins. This is useful in refolding, impurity, and post-translational modification assays on intact biogeneric proteins.

When one looks at peptide mapping applications, performance requirements are significantly different versus intact protein separations, as increased retention and selectivity are required to separate the large number of peptides generated in peptide mapping applications. Because increased resolution is a higher priority versus speed, a larger particle (3.6 μm) 100 Å HPLC/UHPLC column (Aeris PEPTIDE) was developed. It is specifically designed to improve separations of peptide and peptide mapping applications by allowing the use of longer columns at lower backpressures. In this application note, the increased resolution that the longer Aeris PEPTIDE 3.6 μm XB-C18 provides will be demonstrated.

A widepore core-shell HPLC/UHPLC column (Aeris WIDEPORE) has been introduced that is specifically designed to improve protein separations. Rather than utilize a similar morphology of small molecule core-shell columns with larger pores, a completely different particle was developed that takes into account the slower diffusion of proteins into porous particles. When one looks at different intact protein separations, the resolution of different glycoforms of therapeutic IgG antibodies probably stands out as one of the more difficult due to the large size and structure of IgG. In this article improved separation of IgG is shown using Aeris coreshell HPLC/UHPLC columns.

Cyclosporine A, tacrolimus, sirolimus, and everolimus are four of the most commonly administered immunosuppressant drugs and play a central role in the success of tissue and organ transplants. These drugs are most typically analyzed from whole blood using LC/MS/MS. However, because of the analytical challenges posed when working with whole blood, many of the published methods rely upon complex and/or expensive extraction steps prior to use of the actual analytical column. This application note presents a rapid and effective method for the analysis of these four immunosuppressants from whole blood that uses a simple protein precipitation step followed by direct injection onto a wide-pore core-shell HPLC column (Aeris WIDEPORE 3.6 μm XB-C18). The method displays excellent accuracy and is sensitive down to the low μg/L (ng/mL) range.

The ever growing interest in protein therapeutics has found researchers looking for improved solutions for identifying posttranslational modifications of proteins. For the analysis of large, intact proteins like recombinant IgG therapeutics, Phenomenex offers two bioseparation product lines (Aeris and BioSep) for improved solutions in identifying low-level post-translational modifications. Aeris WIDEPORE is a reversed phase core-shell HPLC/UHPLC column that features improved resolution and recovery versus traditional fully porous 300 Å media. BioSep columns offer high performance gel filtration separations that offer improved resolution and inertness versus other aqueous GFC columns. This application note features examples of the use of each column for analyzing recombinant IgG.