Met oxidation to sulfoxide
Air, DMSO, peroxide, even residual H₂O₂ from HPLC silica oxidize Met thioether to sulfoxide. +16 Da. Almost universal for Met-containing peptides without active prevention.
Why it happens (mechanism)
S(II) of the thioether is nucleophilic; any electrophilic O source (singlet O₂, H₂O₂, DMSO under acid, performate) gives Met(O). Sulfoxide can be re-oxidized to sulfone (+32) under harsher conditions. Sulfoxide is generally biologically inactive at the Met position.
When it strikes (triggers)
Any aerobic exposure. DMSO-containing buffers (used for disulfide formation). Performic acid (intentional or contaminant). HPLC mobile phase with persulfate or peroxide. Also: TFA cleavage in the presence of certain scavengers (DMSO, NCS) accelerates it.
How to spot it (MS signature)
+15.99 Da (matches Trp/His/Tyr/Cys oxidation isobarically — distinguish by which residue is in the sequence and by MS/MS — Met(O) loses CH₃SOH at -64 readily).
How to prevent it
- Use NH₄I + DMS (or DMSO in small amounts as reductant cocktail) in TFA cleavage to keep Met reduced. Standard scavenger cocktail K = 82.5/5/5/5/2.5 TFA/phenol/H₂O/thioanisole/EDT.
- Replace Fmoc-Met-OH with Fmoc-Met(O)-OH (sulfoxide protection), then reduce post-cleavage with NH₄I or TMSBr — keeps Met masked through synthesis.
- Lyophilize from acidic solution; store under inert atmosphere (Ar/N₂) at -20 °C.
- For HPLC: degas mobile phase, avoid old/peroxide-containing TFA stock.
If it already happened (salvage)
- Reducible: NH₄I in TFA (5–30 min, room temp) cleanly reduces Met(O) → Met. Or TMS-Br / DMS at low temp. This is one of the few oxidations that's routinely reversible.
Source
Yi Yang, Side Reactions in Peptide Synthesis (Elsevier, 2016), Chapter 9, §9.2.