Mass spectrometry: how a peptide’s identity is confirmed

Purity tells you how clean a lot is. Identity tells you whether the compound is the one on the label. On a certificate of analysis (COA), identity is confirmed by mass spectrometry.

What it measures

Every peptide has a molecular mass set by its amino acid sequence. The mass spectrometer ionizes the molecules in the sample and measures their mass-to-charge ratio. The result is a spectrum with peaks at specific values.

Expected mass and observed mass

The lab compares two numbers:

Theoretical mass. Calculated from the formula of the compound.
Observed mass. What the instrument measured in the sample from the lot.

If the two agree within the tolerance of the method, identity is confirmed. A larger difference means the sample is not the expected compound, or that it has been modified.

Why there are several peaks

One molecule can pick up more than one charge during the analysis. A peptide spectrum therefore often shows several peaks that correspond to the same mass at different charge states. The report states which species each peak belongs to. Several peaks do not mean several compounds.

What to check on the COA

The compound name and lot number match the ones on your vial.
The report shows the theoretical and the observed mass, not just the word “conforms”.
The spectrum is included, with the analysis date and the name of the lab.

Identity and purity are read together

Mass spectrometry confirms what the compound is. HPLC shows how much of the sample is that compound. A reliable lot passes both tests, and both appear on the same COA under the same lot number.

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