In a previous post, I evaluated how flow rate can impact my purification efficiency using flash chromatography. I noticed though, at high flow rates a significantly later elution time for my peptide. I hypothesized that the increased pressure was driving the compound further into the pores, increasing the overall interaction with the stationary phase and causing the increased retention. We know that the particle size and particle pore size impact resolution and purification efficiency, so how does flow rate play a role with a different stationary phase?
In today’s post I’ll evaluate several flow rates using a reversed phase stationary phase material with slightly larger diameter particles that possess significantly smaller pores. The smaller pores should limit the access of the peptides to the stationary phase and negatively impact the purification.
Continue reading How does flow rate affect my peptide purification efficiency when using a small pore stationary phase?
Wow! I can’t believe it’s been a year since this whole blogging adventure started. This started as a technical resource for me and my colleagues within the peptide synthesis and purification space. The idea was to enable easy access to some of the hard-to-find answers for common questions that I have encountered working with different peptide groups.
The blog has grown steadily this year and for that I thank you. In the following post, I’ll highlight a few of the most popular posts from the past year as well a few of my favorites. I hope you will consider providing feedback using the survey link so that we can continue to grow, keeping the content interesting and relevant.
Continue reading Celebrating one year of the peptide synthesis and purification blog!
I’ve recently worked with several peptide groups that are trying out flash purification with their peptides for the first time. And it never fails, every single interaction includes the question “what flow rate should I use for these cartridges?”
There is a lot of information available highlighting optimal flow rates for HPLC method development, but very little information for larger particle stationary phases. I personally have used a wide range of flow rates for my peptide purification with differing outcomes. So in today’s post I’d like to have a more thorough discussion about mobile phase flow rate and it’s impact on your chromatography.
Continue reading What mobile phase flow rate should I use for my 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.
Continue reading Can I improve my peptide purification by increasing the column length?
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.
Continue reading How much peptide is recovered from a reversed phase C18 cartridge during flash purification?