Mass spectrometry in peptide validation
How mass spectrometry confirms the identity of a synthetic peptide by matching the observed mass to the theoretical mass of its sequence.
Where HPLC tells you how much of a sample is the main component, mass spectrometry tells you what that component is. It is the identity check on a peptide Certificate of Analysis.
Why mass confirms identity
Every peptide sequence has a precisely calculable mass. Measuring the observed mass of a synthesised sample and comparing it to the theoretical mass of the intended sequence confirms that the right molecule was made — within the instrument's tolerance. A mismatch points to a deletion, an incomplete deprotection, or an unintended modification.
MALDI-TOF and ESI
Two ionisation methods dominate peptide work. MALDI-TOF (matrix-assisted laser desorption/ionisation, time-of-flight) is fast and tolerant of salts, giving a clear singly-charged ion for most peptides. ESI (electrospray ionisation) produces multiply-charged ions and pairs naturally with liquid chromatography for LC-MS. The COA should name which was used.
Interpreting the result
Expect the observed monoisotopic or average mass to agree with the calculated value within a few mass units (method-dependent). Read mass spectrometry together with the HPLC purity figure and the counter-ion analysis: identity, purity and salt content are three different questions, each answered by a different measurement.
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