Mechanically, the honest answer is that the achievable range of plausible purity figures for a given vial is wider than most people expect.
Gradient slope controls resolution, and gentler slopes resolve co-eluting impurities into separate peaks — so the better method reports the worse purity number.
Mass shifts and what they usually mean
| Δ mass (Da) | Most likely cause | Distinguishing feature |
|---|
| +1 | Deamidation (Asn or Gln) | New peak, slightly earlier retention |
| −17 | Loss of ammonia | Often with deamidation |
| −18 | Dehydration / succinimide | pH-dependent, reversible |
| +16 | Oxidation (Met, Trp) | Earlier retention, light-related |
| −128 | Missing Gln or Lys | Deletion sequence from synthesis |
| 0 | Isomer: racemisation or scrambling | Same mass, shifted retention |
In practice, mobile phase additive choice affects ionisation and peak shape — TFA gives sharp peaks but suppresses mass spectrometry signal, formic acid gives worse peaks but preserves signal.
The Arrhenius relationship for peptide degradation is the basis of accelerated stability testing and also governs how quickly methods drift with temperature.
The practical summary: ask for the chromatogram and the method, and ignore the headline number until you have both.
6Adding for future readers: the certificate should carry the lot number, not just a batch code. – tare_weight 8 months ago add a comment