How to read a peptide COA: HPLC, mass spec and batch checks
Why short peptides are different
A seven-residue peptide like Semax or Selank is small enough that mass spectrometry gives an unambiguous identity check, and simple enough that HPLC should show one dominant peak. That makes a poor COA easy to spot. It also means a clean-looking number is not enough: purity, identity and quantity are three separate questions.
HPLC purity
Purity is the area of the main peak as a percentage of all integrated peaks, usually detected by UV at 214 or 220 nm. Open the chromatogram instead of reading only the summary. Look for a sharp main peak, a flat baseline, and whether small impurity peaks were integrated or ignored. Purity says nothing about whether the main peak is the right molecule.
Mass spectrometry and identity
Identity comes from the measured mass. For electrospray ionisation, the singly protonated ion [M+H]+ is the usual signal, calculated from the sequence as:
- Semax (MEHFPGP): monoisotopic 813.35 Da, so [M+H]+ ≈ 814.36 (average mass 813.9).
- Selank (TKPRPGP): monoisotopic 751.43 Da, so [M+H]+ ≈ 752.44 (average mass 751.9).
Because Selank contains lysine and arginine, doubly charged ions ([M+2H]2+) are also common, so a spectrum showing a signal near half the mass plus one proton is expected rather than a flaw. Semax contains a methionine residue, which can oxidise to a sulfoxide (+16 Da), so a small +16 signal is something to look for on a COA and in storage.
Peptide content vs purity
Lyophilised peptides contain counter-ions and residual water, so the fraction of the vial that is actually peptide (net peptide content) is lower than the HPLC purity figure. A COA that reports measured quantity per vial answers a different question from one that reports purity only. If a report gives only a purity percentage, treat the labelled amount as unverified. The sensitivity of short peptides to enzymes in biological media, noted in the Semax literature,1 is a reason to follow the supplier’s storage guidance for dissolved material.
Traceability and verification
- The batch number on the report must match the vial and packaging.
- The testing laboratory should be independent of the seller.
- A verification key or URL on the lab’s own site lets you confirm the report was not edited. Check it, do not rely on the PDF image alone.
- The test date should plausibly precede the sale of that batch.
Null Bio Lab publishes independent reports with verification links on the COA page; the current Semax batch and its report are on the Semax product page.
Red flags
- A purity figure with no chromatogram.
- No mass data, or a mass that does not match the calculated value above.
- A report from the seller’s own “in-house” lab only.
- A batch number on the COA that differs from the vial.
- Round numbers repeated across different compounds.
Related: Semax & Selank research guide · Semax vs Selank.
Frequently asked questions
What should a peptide COA include?
HPLC purity with the chromatogram, a measured mass that matches the calculated value, measured quantity, a batch number matching the vial, and a verification key on the testing lab’s own site.
Is HPLC purity enough?
No. Purity says how much of the signal is one peak, not that the peak is the right molecule. Identity needs a mass-spectrometry result.
Why is net peptide content lower than purity?
Lyophilised peptide contains counter-ions and residual water, so the peptide fraction of the vial is lower than the HPLC purity figure.
How do I verify a report?
Enter the report’s key on the independent laboratory’s own verification page instead of relying on a PDF image.
References
- Potaman VN, et al. Degradation of ACTH/MSH(4–10) and its synthetic analog semax by rat serum enzymes: an inhibitor study. Peptides. 1993. doi:10.1016/0196-9781(93)90137-6.
Related reading
For newly published papers on this and other peptides, see the live feed on the Research Desk.