One common technique in HPLC for improving difficult peptide separations is to extend the column length, a topic I explored for flash chromatography in a previous post. However, alternative purification strategies are sometimes necessary as the purification bottleneck grows with increasing peptide library size, both in number and scale.
In this post, I explore using two identical size cartridges in series with each packed with a different stationary phase. I wanted to try this to see if I could improve peptide purity with the ultimate goal of reducing the time demand of peptide purification. Continue reading Using mixed stationary phases to improve your peptide purification with flash chromatography
There are several strategies often employed to improve peptide purity achieved using reversed phase HPLC. These strategies can include, changing column length, particle size, particle functionality (C4 vs C18). I have experimented a bit with some of these criteria while purifying peptides using reversed phase flash chromatography but one obvious change that I have not yet explored is the length of column.
In today’s post, I’ll explore how the length of the cartridge affects the overall resolution and purification efficiency using reversed phase flash column chromatography.
In peptide purification, sample loading onto the column is rarely considered. Most, if not all, HPLC instruments come equipped with a sample injection loop which demands a liquid injection of the sample for purification. If you decide to use flash chromatography to purify your peptides though, liquid injection is no longer the exclusive method for sample introduction to the column. Alternatively, dry loading crude material is a strategy often used in small molecule purifications, particularly when sample solubility concerns arise.
The first question I asked myself when considering a new sample loading strategy is whether or not the purification efficiency will be maintained. A close second though is whether or not the loading method will cause significant differences in peptide recovery.
In today’s post, I’ll compare recovery efficiencies for peptides purified using reversed phase flash chromatography but loaded onto the cartridge using either direct liquid injection or dry loaded onto reversed phase material. Continue reading Does loading method influence my peptide recovery after purification?
Whether it’s the bonded stationary phase, particle size, or even particle pore size, scientists today are offered a plethora of choices when it comes to reversed phase HPLC columns. An often acknowledged concern in the peptide community though is peptide recovery from reversed phase purification efforts, particularly for precious peptide mixtures. But how is peptide recovery impacted when you use reversed phase flash chromatography for purification?
In today’s post, I’ll compare recovery levels for two peptides that differ in length as well as crude purity using reversed phase flash chromatography. In addition to comparing two peptides, I’ll also evaluate how recovery is impacted by altering the mobile phase pH.
Big pharmaceutical companies have begun to refocus their efforts towards peptide discovery projects with the hopes of identifying the next big peptide drug. There are often hundreds to thousands of peptides synthesized as part of these efforts, demanding parallel synthesis platforms and room temperature peptide synthesis protocols.
Previously, I identified a minimum number of amino acids equivalents required to ensure a high quality microwave synthesis. Conducting synthesis at room temperature will certainly require different conditions than microwave heating. Let’s explore how the number of equivalents will impact the synthesis results.
In a previous post, I did some work evaluating the efficiency of alloc removal with tetrakis palladium using microwave assistance and atmospheric conditions, which worked beautifully. Given the known sensitivity of palladium catalysts (see Derek Lowe’s post for a humorous dialogue), I sought to further explore the sensitivity of palladium towards the alloc removal in the context of a peptide.
In this post, I’ll explore a variety of atmospheric, room temperature alloc deprotection conditions aimed at evaluating the catalytic lifetime of palladium tetrakis for effective alloc removal.