Tailing peaks can be a problem when we are doing liquid chromatography (LC), and secondary silanol interactions are one of the causes. Here, using the example of basic drugs, we explain what causes them and cover a few steps that we can take to minimize the effect of these secondary silanols on our peak shape when we are troubleshooting.
- All of My Peaks are Tailing! What Should I Do?
- Troubleshooting HPLC – Tailing Peaks
- When Should You Use a Buffer for HPLC?
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Thanks for joining the latest Restek Tip, where I'll discuss troubleshooting peak tailing and LC analysis, and some ways to actually overcome this. We all know that peak tailing can be due to a number of different factors, including secondary silanol interactions, contamination, column overloading... the list really can go on and on. I really want to focus on secondary silanol interactions and how we can mitigate them during our development phase. One thing that I do when I start to look at peak tailing and what are some things that I can do to help reduce peak tailing, is really look at all of the analytical conditions that I have set up in my method.
Once I've gone over the mobile phases that I've chosen, the gradient, temperatures, etc., I'll start to dig a little bit deeper, and look at analyte structures. If I look at analytes, and think about positively charged compounds, anything that has maybe an amine group, like a small basic drug, that positive charge of the drug can actually start to interact with the negative charge of residual silanols on the surface of the silica.
That interaction is what's actually creating peak tailing. One way to actually get around this is to add buffer to your mobile phases. In typical LC-MS analysis, we're already adding acid to our aqueous and organic components of our mobile phases. The addition of complementary salts, like ammonium formate to formic acid, can really help reduce some of those silanol interactions.
Essentially, what happens is, the positive charge of the buffer salt that we're adding can interact with the negative charge of the silanol surface on the silica, so that's going to drive that interaction and really not allow your analyte to interact with the surface of the silica. The primary retention mechanism is going to be driven by the stationary phase you've chosen. One thing that's important to remember is to include the buffers in both your aqueous and organic components as you're running gradient analyses as to mitigate the tailing that you see in both early and late eluting peaks.
So, the next time you're looking at some small basic drugs and having trouble with peak tailing, try adding some buffer to your mobile phase, to help get rid of that.
Thanks for joining the latest Restek Tip.