Calculated, not measured
The theoretical molecular weight follows from the sequence: sum the residue masses and subtract a water molecule for each bond formed. Mass spectrometry then measures what is actually present, and agreement between the two is the identity check.
Average and monoisotopic mass
Elements exist as isotopes. Average mass uses the natural abundance of each; monoisotopic mass uses only the most common isotope. For small peptides the two differ by a fraction of a percent — enough to matter when comparing a measurement against a specification, which is why a COA states which one it quotes.
Why size changes everything else
A 340 Da tripeptide and a 22,000 Da protein sit at opposite ends of the catalogue. Size determines whether a molecule can be chemically synthesised, how it is purified, how fragile it is, and how precisely its mass can be confirmed.
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