HPLC vs mass spec: what each peptide test actually proves

Most peptide certificates show a purity figure, often 98% or 99%. That number usually comes from HPLC. It is useful, but on its own it can be misleading.

What HPLC does

High-performance liquid chromatography pushes a dissolved sample through a column. Different molecules come out at different times and show up as peaks on a chart. The purity figure is the size of the main peak compared with all the peaks. HPLC tells you the sample is mostly one thing. It does not, by itself, tell you what that thing is.

What mass spectrometry does

Mass spectrometry measures the mass of the molecules in the sample. Each peptide has a known mass, so a match between the measured and expected mass confirms identity. Labs often combine the two (LC-MS) so the main HPLC peak is identified directly.

To find the expected mass to compare against a COA, you can use the peptide molecular weight calculator from Codex Research Lab.

Why you want both

A vial can be 99% pure and the wrong peptide. A vial can be the right peptide at poor purity. Only the two tests together answer both questions. Neither one measures endotoxins or how much peptide is in the vial, which need separate tests.

For the full checklist, see how to read a COA.