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      How to choose your acidic mobile phase modifier for peptide purification using Reversed-Phase flash chromatography

      Jun 17, 2020 1:32:27 PM / by Elizabeth Denton posted in Troubleshooting and Optimization, Sfär, Peptide Purification

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      When purifying crude peptides, we always use a mobile phase modifier. The modifier simplifies a potentially complex purification by driving the desired peptide, and any impurities in the sample, into a single protonation state. That way, you're only trying to purify a single peptide, rather than many variants of the same peptide that only differ by the presence or absence of a couple protons - from many peaks to one!

      Acids are commonly used as mobile phase modifiers. Some groups choose trifluoracetic acid (TFA), while others choose formic acid (FA). But are there others that should be considered as well? In today's post, I'll explore the use of an acidic buffer as a mobile phase modifier and compare the purification efficiency to that observed with TFA.

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      How does mobile phase modifier concentration impact peptide purity with flash chromatography?

      Jun 2, 2020 6:44:01 PM / by Elizabeth Denton posted in Reversed-phase, Troubleshooting and Optimization, Sfär, Peptide Purification

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      We all use mobile phase modifiers, regardless of purification strategy, for purification of our synthetic peptides.  With ionizable side chains, something that affects the pH of the mobile phase is a must to ensure that the desired peptide (and any impurities) elute from the column in a single, well defined peak.  

      While a 0.1% modifier concentration is basically standard these days (sometimes you'll see different),  I started to wonder if this was really enough modifier to fully protonate all possible side chains when using flash chromatography to purify peptides.  In today's post I'll look into the effects of different mobile phase modifier concentrations with respect to general peak shape and final sample purity.

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