Mass Spectrometry

Mass spectrometry (MS) ionises a sample and records mass-to-charge ratios, from which the molecular mass is derived. On a peptide certificate it settles the identity question that chromatography leaves open: does this vial contain the sequence printed on the label? Electrospray ionisation (ESI-MS) and MALDI-TOF are the methods most often quoted.

Interpreting the MS result

Set the measured mass against the calculated mass derived from the sequence. The two should match within the tolerance stated for the instrument — usually a fraction of a dalton on a current ESI system, a few daltons on MALDI-TOF. Peptide ESI spectra display a set of multiply charged ions ([M+2H]2+, [M+3H]3+) which software converts into one neutral mass; a certificate listing only a raw m/z value without the charge state is incomplete.

Telling mass shifts

Typical offsets carry meaning: +16 Da points to oxidation, most often at methionine; −0.98 Da separates a C-terminal amide from the free acid; +42 Da signals acetylation; +1 Da may indicate deamidation. A discrepancy equal to exactly one residue mass suggests a missing or swapped amino acid.

Identity plus purity

An excellent HPLC value is meaningless if the molecule is the wrong one. Identity together with purity, measured on the same lot, is the minimum credible combination on a certificate of analysis.

Related terms

See mass spectrometry and peptide identity.